Distribution of phytoplasma diseases in the Lombardy poplar tree population of Zagreb urban area

Phytoplasmas are uncultivable prokaryotic wall-less pathogens belonging to the class Mollicutes that inhabit plant phloem and insects. Their identification and classification is difficult and mainly based on the polymorphism of their 16S rRNAgene sequences. Aster yellows (AY; \u27Candidatus Phytoplasma asteris\u27) phytoplasmas (ribosomal subgroup 16SrI-P) had been previously identified in several Populus nigra L. \u27Italica\u27 (Lombardy poplar) trees from the urban area of Zagreb. The aim of this research was to examine phytoplasmosis distribution in the poplar tree population of the wider Zagreb urban area. Total nucleic acids were extracted from leaf samples of 30 symptomatic and 4 asymptomatic trees. Phytoplasma 16S rDNAwas amplified in direct and nested PCRs by using universal and group-specific primers. The pathogens were classified on the basis of 16S rDNA amplicon RFLP analyses. Phytoplasmas belonging to the 16SrI ribosomal group (AY) were detected in 12 out of 34 trees examined. In addition, a phytoplasma putative gene for aa kinase was analyzed for positive samples. RFLP profiles from 10 samples were referable to the phytoplasma 16SrI-P ribosomal subgroup previously found only in poplars from Zagreb area. In two samples, unique restriction patterns were found, showing the presence of molecular variability within this conserved gene region. In the northern and north-western part of the area the infection was equally distributed, while in the southern part of the city phytoplasmas were not detected in sampled trees. Further research, including a search for potential insect vectors, is needed in order to indentify the ecological and epidemiological implications of these diseases and their impact on the sanitary status of ornamental trees in the city of Zagreb

Hidden genetic diversity of ‘Candidatus Phytoplasma solani’ strains in Istrian vineyards: how small can be rich

Grapevine Yellows (GY) are economically important diseases associated to phytoplasmas (genus ‘Candidatus Phytoplasma’), uncultivable bacteria from the class Mollicutes. In Euro-mediterranean region, GY are mainly attributed to phytoplasmas belonging to ribosomal 16SrXII-A subgroup ('Ca. P. solani'; Bois Noir phytoplasma; BNp) and 16SrV group (flavescence dorée phytoplasma; FDp), with the latter one being a quarantine pathogen (Plavec et al. 2015, Quaglino et al. 2013). Although symptomatology can be similar, epidemiology of FDp and BNp is very different as they are transmitted by different insect vectors and could have different alternative hosts. In Croatia, GY diseases have been continuously monitored since 1997 with FDp appearing only in 2009 (Šeruga Musić et al. 2011). While BNp is more diverse and widespread, FDp belonging to 3 map phylogenetic clusters are differently distributed throughout regions. Istria is heavily affected with FDp since 2014 with only one genotype (mapFD2/M54) being attributed to great losses (Plavec et al. 2019). On the contrary, information on BNp diversity is scarce with only couple of genotypes detected so far (Plavec et al, manuscript in preparation). Therefore, the aim of this case study was to assess the diversity of BNp genotypes in a small selected area of Istria, in order to draw attention to the presence and importance of BNp as GY agent in this important Croatian viticultural region

Geographical distribution of 'flavescence dorée' phytoplasmas in Croatian grapevines

Abstract Seven Croatian grapevine samples found positive for the presence of 'flavescence dorée'(FD) phytoplasmas in 2010 survey confirm the first findings of FD phytoplasma from previous year, as well as the existence of new grapevine yellows heavily affected narrow area west and southwest of Zagreb along the border with Slovenia. Two new locations 100 km east and 60 km north-east of Zagreb, respectively, were found to be affected, widening the potential FD-risk zone and placing the capital in its centre. Grapevine varieties infected with FD-related phytoplasmas encompassed widely known Pinot Noir and Riesling, but also indigenous red Plemenka Crvena and Ružica Crvena, as well as white variety Škrlet. Preliminary molecular characterization results suggest the variability of 16SrV-C identified phytoplasmas, especially from the locations west and southwest of Zagreb

Distribution and epidemiology of Flavescence doree phytoplasma in Croatia

Žutice vinove loze nanose velike štete jer smanjuju prinos i kvalitetu grozda to naposlijetku dovode do potpunog propadanja vinove loze. Uzročnik zlatne žutice vinove loze, fitopatogena bakterija Flavescence dorėe fitoplazma (FD), u Europi se ubraja u karantenske patogene, a rasprostranjuje se cikadom Scaphoideus titanus Ball. U posljednjih desetak godina sustavno se prati prisutnost i proširenost fitoplazmoza vinove loze programom posebnog nadzora, a 2009. prvi je put utvrđena prisutnost fitoplazme FD na nekoliko lokacija. Od tada do 2012. porastao je broj pozitivnih nalaza i broj lokaliteta na kojima je utvrđena fitoplazma FD. S ciljem da se utvrdi izvor i način njezina rasprostranjenja za studiju epidemiologije odabrana su dva lokaliteta: Brckovština (Križevci) i Jagnjić Dol (Samobor). Fitoplazma FD potvrđena je u 8 od 10 prikupljenih uzoraka vinove loze te u 12 od 21 uzoraka vektora S. titanus. Molekularnim analizama utvrđeno je da su na tim lokacijama prisutne različite skupine fitoplazme FD: skupina FD-1 u Brckovštini te skupina FD-3 u Jagnjić Dolu.Grapevine yellows (GY) diseases cause severe economic damage by reducing yields and quality of grape and ultimately lead to overall decline of plants. One of most important GY agents, Flavescence doree (FD) phytoplasma, is a quarantine organism in the Europe due to its Cicadelidae vector Scaphoideus titanus Ball. which disseminates FD in epidemiological manner. GY diseases have been continuously monitored during the last ten years throughout the official annual survey programme and for the first time in the 2009. the presence of FD phytoplasma was confirmed at several locations. From the first fmding the number of FD infected samples as well as the number of affected locations has increased. With the aim to elucidate FD phytoplasma epidemiology in Croatia and establish source and ways of dissemination, two locations - Brckovstina (Kri2evci) and Jagnjic Dol (Samobor), representing FD phytoplasma foci have been chosen. FD phytoplasma presence was confirmed in 8 out of 10 grapevines and in 12 out of 21 S. titanus samples. Molecular analyses clearly showed a presence of distinct isolate at each of the two locations; the isolate FD-1 at Brckovstina and FD-3 at Jagnjic Dol

First viruses infecting cockspur coral tree (Erythrina crista-galli L.): discovery of a putative novel capillovirus and a new host for the prune dwarf virus

peer reviewedA cockspur coral tree (Erythrina crista-galli, Fabaceae) from the collection of woody ornamentals of the Botanical Garden (Faculty of Science) in Zagreb showed conspicuous virus-like symptoms. The leaf tissue was analyzed by high throughput sequencing (HTS), revealing the presence of two viruses: prune dwarf virus (PDV, genus Ilarvirus) and an unknown virus belonging to the genus Capillovirus (family Betaflexiviridae). The complete sequence of PDV RNA3 of 2,129 nucleotides (nts) coding for the coat and movement proteins was obtained. The complete coding region spanning 6,483 nts was obtained for the capillovirus. It contained all expected open reading frames, and its maximum nucleotide identity was 42% with apple stem grooving capillovirus sequence (GenBank accession number LC143387). As this is far below the threshold for species delineation within the genus Capillovirus, we propose the name Erythrina capillovirus (ErCV) for this putatively new virus and a new virus species Capillovirus ErCV in the family Betaflexiviridae. RT-PCR confirms the presence of ErCV and PDV in symptomatic E. crista-galli, an unusual and exotic fabaceous host with no viruses recorded yet. Asymptomatic Erythrina plants were tested negative for the two viruses in RT-PCR, which, together with their presence in the symptomatic plant, suggests the capillovirus and/or PDV might play a role in eliciting observed symptoms and deserve further investigations

Euscelis incisus in Bois noir-affected vineyards of Istria (Croatia): vector competence and syntopic occurrence with E. lineolatus

Bois noir (BN) disease of grapevine ('Ca. Phytoplasma solani', CaPsol,16S rRNA XII-A) was first molecularly confirmed to affect vineyards in Croatia more than two decades ago (Škorić et al., 1998). Presently, BN is widespread, occurring in most of the wine regions of the country (Plavec et al., 2018). The diversity of CaPsol genotypes in Croatian vineyards indicates complex epidemiology, which was recently confirmed in Istria, the westernmost grape-growing county of Croatia in the northeastern Adriatic Sea (Delač et al., 2023). Euscelis incisus (Kirschbaum, 1858) is a nitrophilous, polyphagous leafhopper, widely distributed throughout the Western Palaearctic region, inhabiting diverse habitats (Nickel, 2003). Due to its wide distribution and ability to inhabit a variety of ecologically distinct environments, this leafhopper can encounter diverse phytoplasmas that invade a wide range of plant species. It has been established as a vector of the 16Sr I-B/C, III-B, and XII-A phytoplasma subgroups and a carrier of the I-F/R, II-E, IX-C/E, and XI-G subgroups (Jakovljević et al., 2020). Its congener, E. lineolatus (Brullé, 1832) was found to harbor I-B/C and XII-A subgroups, while its ability to act as a phytoplasma vector has not been confirmed so far (Landi et al., 2013). The aim of this study was to investigate the epidemiological and vector roles of E. incisus and co-occurring E. lineolatus in transmitting CaPsol in BN-affected vineyards of Istria

Legacy of Plant Virology in Croatia—From Virus Identification to Molecular Epidemiology, Evolution, Genomics and Beyond

This paper showcases the development of plant virology in Croatia at the University of Zagreb, Faculty of Science, from its beginning in the 1950s until today, more than 70 years later. The main achievements of the previous and current group members are highlighted according to various research topics and fields. Expectedly, some of those accomplishments remained within the field of plant virology, but others make part of a much-extended research spectrum exploring subviral pathogens, prokaryotic plant pathogens, fungi and their viruses, as well as their interactions within ecosystems. Thus, the legacy of plant virology in Croatia continues to contribute to the state of the art of microbiology far beyond virology. Research problems pertinent for directing the future research endeavors are also proposed in this review

Preliminary data on the natural reservoir plants and insect carriers of Flavescence dorée and related phytoplasmas in Croatia

The occurrence of Flavescence dorée (FD) disease of grapevine, its associated phytoplasma, FDp (a member of the 16SrV taxonomic group of elm yellows, subgroups V-C and -D), and the main insect vector, the deltocephalinae leafhopper of North American origin, Scaphoideus titanus, was first evidenced in vineyards in Croatia in 2009 (Šeruga Musić et al., 2011; Plavec et al., 2015). Contemporary identification of FDp in the climbing shrub Clematis vitalba as one of the main natural reservoir plants in North Italy and the Balkans was also confirmed for Croatia (Filippin et al., 2009). Subsequent studies aimed at determining the genetic diversity and structure of FDp populations and tracing transmission pathways in Croatia revealed the invasive tree Ailanthus altissima and the natural riparian tree Alnus glutinosa as natural FDp reservoirs (Plavec et al., 2019). However, the roles and epidemiological importance of each of the identified reservoir plants remained unclear due to the limited number of elaborated samples. Regarding the natural insect vectors of FDp from reservoir plants to grapevine and their association with natural reservoir plants on the wider territory of Croatia, particularly beyond vineyards in their natural habitats, not much is known. To understand the epidemiological importance of FDp natural plant hosts and insect vectors, we performed surveys targeting clematis, alders, and other tentative natural woody hosts of FDp in vineyard surroundings and natural habitats in wine-producing regions of Croatia

Comparison of clinical and sewage isolates of Acinetobacter baumannii from two long-term care facilities in Zagreb; mechanisms and routes of spread

In the previous studies OXA-23-like and OXA-24-like β-lactamase were reported among Acinetobacter baumannii in both hospitals and long-term care facilities (LTCF) in Croatia. The aim of this study was to analyze clinical and sewage A. baumannii isolates from two nursing homes in Zagreb, with regard to antibiotic susceptibility and resistance mechanisms, to determine the route of spread of carbapenem-resistant isolates. Nine clinical isolates were collected from February to May 2017 whereas in April 2017, ten A. baumannii isolates were collected from sewage of two nursing homes in Zagreb. Antibiotics susceptibility was determined by broth microdilution method. The presence of carbapenemase and extended spectrum β-lactamases (ESBL) encoding genes was explored by PCR. Conjugation and transformation experiments were performed as previously described. Genotyping was performed by SG determination, PFGE and MLST. Seven clinical isolates were positive for blaOXA24-like whereas two clinical and environmental carbapenem-resistant isolates, respectively, were found to possess blaOXA-23-like genes. Attempts to transfer imipenem resistance were unsuccessful indicating chromosomal location of blaOXA-23 gene. All carbapenem-resistant isolates belonged to SG- 1 (IC-2) whereas the rest of the isolates susceptible to carbapenems were allocated to SG- 2 (IC-1). PFGE analysis revealed low degree of genetic variability within both IC- I and IC- II. MLST corroborated that two environmental OXA-23 isolates belong to the ST-195. This study showed dissemination of OXA-23 producing A. baumannii from the nursing home into the urban sewage. Disinfection of nursing home sewage should be recommended in order to prevent the spread of resistance genes into the community sewage